Abstract The global burden of malaria and the widespread drug resistance in Plasmodium falciparum demand novel and improved therapy. The availability of large libraries of small molecular compounds provides a rich resource for the discovery of antimalarial drug candidates. Screening of these structurally diverse small-molecule libraries requires the development of a robust, high-throughput, and highly reproducible assay. Luminescence-based assays are highly preferred and widely used in high-throughput screening (HTS) in many biological systems as they have better signal-to-noise ratios, are as sensitive as radiolabel-based assays and easy to automate. We propose to develop a malaria parasite cell-based assay using the firefly luciferase reporter system. So far, we have developed a luciferase-expressing P. falciparum line and optimized a luciferase assay for 96-well microtiter plates. In the first phase of the project, we will validate and optimize this assay in a 384-well format and develop another luciferase- expressing line for complementary drug screening. In the second phase, we propose to perform a pilot HTS of a preexisting 3,000-compound library representing a structurally- diverse 100,000 compound library. Positive hits from the primary screening will be confirmed by dose-ranging experiments using the standard [H3] hypoxanthine incorporation assay. This high-throughput luminescence-based assay system will be a very useful tool for future HTS efforts in search for novel antimalarial therapies. Narrative With an annual death toll exceeding one million, malaria is a significant public health problem in many developing countries. With the emergence and spread of parasites resistant to many antimalarial drugs, there is a pressing need for the discovery of novel and improved drugs for malaria control. To facilitate antimalarial drug discovery, we propose to develop a highly sensitive, cell-based, one-step luminescent drug screen assay for high throughput screening of large chemical libraries. [unreadable] [unreadable] [unreadable]